Chlamydia pneumoniae is an intracellular bacterium that causes a spectrum of respiratory infections and is associated with cardiovascular atherosclerotic disease. Although most studies on C. pneumoniae have focused on providing evidence for its pathogenic role in atherosclerosis, knowledge of the chlamydial antigens and immune mechanisms that lead to protective or adverse immune responses remains limited. Because CD8+ T cells play a critical role in defense against most intracellular pathogens, we hypothesize that C. pneumoniae infection in mice primes a multispecific CD8+ CTL response capable of inhibiting chlamydial growth and that CTL epitope-based immunization strategies can reduce bacterial burdens in the lungs of challenged mice. We will test this hypothesis through the following aims: 1) Identify C. pneumoniae CD8+ CTL target antigens and define the mechanisms by which CD8+ T cells inhibit bacterial growth. We will identify antigens of the murine CD8+ CTL response to C. pneumoniae using MHC class I binding motif-bearing synthetic peptides from putative chlamydial CTL target antigens and mass spectrometric analyses of MHC class I-bound peptides from bacteria-infected cells. Lung and splenic epitope-specific CD8+ CTL will be tested for their capacity to inhibit development of chlamydial inclusions in infected cells through secretion of type 1 cytokines and lytic /apoptotic molecules. We will also use real-time PCR and immunohistochemistry to detect effector molecules in lung tissue. Effector mechanisms will be evaluated in vivo by assessing bacterial loads in lungs of C. pneumoniae-challenged mice after adoptive transfer of CD8+ CTL lines genetically deficient of effector individual molecules. 2) Characterize the response kinetics of C. pneumoniae epitope-specific CD8+ CTL during infection. We will study primary and recall CD8+ T cell responses in the lungs and spleens of C. pneumoniae-infected mice by ELISPOT, confocal microscopy, and flow cytometry with activation and memory cell markers, and MHC class I tetramers. 3) Determine the capacity of CD8+ CTL-based immunotherapies to prevent or ameliorate C. pneumoniae infection. Selected CTL epitopes will be used to generate peptide- and DNA-based vaccines that will be tested for their capacity to reduce bacterial burden in the lungs of C. pneumoniae-challenged mice. These studies will enhance our understanding of the role of CD8+ T cells in protection against C. pneumoniae and will suggest methods of immunological intervention in infections caused by C. pneumoniae and other chlamydial pathogens.